The process by which T cells are activated, and the consequences of activation (e.g. new gene transcription, cytokine production, apoptosis), are being investigated by a variety of approaches: 1) activation-induced apoptosis of T cell hybridomas is mediated by upregulation of the ligand for Fas (FasL). We have identified a single major site 5' of the transcription initiation site that is required for initiation of fasL transcription. This site is bound by the Egr family of transcription factors, and Egr-2 and Egr-3 (but not Egr-1) are responsible for transactivation of gene transcription at this site. 2) We have found that corticosteroids prevent activation-induced apoptosis, and have hypothesized that this phenomenon regulates antigen-specific thymocyte selection. We have crossed transgenic mice that express antisense glucocorticoid receptor in immature thymocytes with lpr autoimmune mice, and have found that TCR Vbeta usage is altered and autoimmunity and lymphadenopathy are greatly diminished. Moreover, B10.BR (H-2k) mice bearing this antisense transgene no longer respond to the antigen pigeon cytochrome c, although they respond normally to more complex antigens such as PPD and alloantigen. These results demonstrate that glucocorticoids, likely those produced in the thymus itself, do in fact regulate positive and negative selection by antagonizing TCR-mediated signals, and therefore shape the peripheral immune repertoire. Studies on the possible role of glucocorticoids in the generation of autoimmunity are ongoing. 3) Inhibitors of proteasome activity block thymocyte apoptosis. We have found that IAPs (inhibitors of apoptosis) are selectively degraded in proteasomes in response to apoptotic stimuli. The IAPs themselves have ubiquitin protein ligase (E3) activity, and are responsible for their own ubiquitination. How this is regulated in vivo is under investigation. 4) Glucocorticoids are immunosuppressive, largely because they interfere with the transcription of many activation-induced genes. We are examining the possible role of a glucocorticoid-induced gene, GILZ, in this activity.